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Abstract

Liver fibrosis is a major cause for increasing mortality worldwide. Preclinical research using animal models is required for the discovery of new anti-fibrotic therapies, but currently relies on endpoint liver histology. In this study, we investigated a cost-effective and portable photoacoustic/ultrasound (PA/US) imaging system as a potential non-invasive alternative. Fibrosis was induced in mice using CCl4 followed by liver imaging and histological analysis. Imaging showed significantly increased PA features with higher frequency signals in fibrotic livers versus healthy livers. This corresponds to more heterogeneous liver structure resulting from collagen deposition and angiogenesis. Importantly, PA response and its frequency were highly correlated with histological parameters. These results demonstrate the preclinical feasibility of the PA imaging approach and applicability of dual PA/US system.

Fig. 5 Correlative analysis of number of photoacoustic pixels (PA) and heterogeneity index (HI) with histological parameters. Panels a and b depict correlations between PA pixels and % intensity of Collagen I and CD31 staining respectively. In addition, panel c shows the correlation between collagen I and CD31. Panels d and e show the correlation between the heterogeneity index (HI) and collagen I and CD31 staining respectively. Finally, panel f depicts the correlation between number of PA pixels and HI. Correlations were assessed using Pearson's correlative analysis. The dot plots for all the correlations were generated using the GraphPad Prism software. ‘R2’ denotes Pearson correlation coefficient and ‘p’ denotes statistical significance.

Fig. 6 Absorption measurements for collagen I. (a) Absorption spectra measured using a Shimadzu UV-2501 spectrophotometer of collagen I / PBS solution in cuvettes. The concentrations were taken as per the average of collagen I concentration in typical fibrotic livers. (b) Plotting of the absorption at 808 nm for the range of concentrations shows proper linear behavior. Assuming 2% of the liver shows collagen scarring and that the total amount of collagen I is concentrated only in these scars, then the approximate absorption coefficient within scars is (0.025 / cm) / 2% = 5 / cm. This is of the same order as the absorption of hemoglobin in functioning blood vessels.